Identification of bacterial microflora in the midgut of the larvae and adult of wild caught Anopheles stephensi: A step toward finding suitable paratransgenesis candidates

To describe the midgut microbial diversity and to find the candidate bacteria for the genetic manipulation for the generation of paratransgenic Anopheline mosquitoes refractory to transmission of malaria, the microbiota of wild larvae and adult Anopheles stephensi mosquito midgut from southern Iran was studied using a conventional cell-free culture technique and analysis of a 16S ribosomal RNA (rRNA) gene sequence library. Forty species in 12 genera including seven Gram-negative Myroides, Chryseobacterium, Aeromonas, Pseudomonas, Klebsiella, Enterobacter and Shewanella and five Gram-positive Exiguobacterium, Enterococcus, Kocuria, Microbacterium and Rhodococcus bacteria were identified in the microbiota of the larvae midgut. Analysis of the adult midgut microbiota revealed presence of 25 Gram-negative species in five genera including Pseudomonas, Alcaligenes, Bordetella, Myroides and Aeromonas. Pseudomonas and Exiguobacterium with a frequency of 51% and 14% at the larval stage and Pseudomonas and Aeromonas with a frequency of 54% and 20% at the adult stage were the most common midgut symbionts. Pseudomonas, Aeromonas and Myroides genera have been isolated from both larvae and adult stages indicating possible trans-stadial transmission from larva to adult stage. Fast growth in cheap media, Gram negative, and being dominantly found in both larvae and adult stages, and presence in other malaria vectors makes Pseudomonas as a proper candidate for paratransgenesis of An. stephensi and other malaria vectors.